The aim of the present study was to evaluate the dietary administration of Bacillus aerophilus KADR3 on enhancing the immunity and disease resistance against Aeromonas hydrophila infection in Labeo rohita. B. aerophilus culture supernatant exhibited antagonistic activity against fish pathogenic bacteria in an agar well diffusion assay. Four fish groups were fed with either of following diets containing different concentrations of B. aerophilus KADR3 (cfu g−1): 0 (control), 1 × 107 (DI), 1 × 108 (DII), and 1 × 109 (DIII) –cfu g−1. Various innate immune parameters were measured at – 3rd and 6th – week post-feeding. At the end of 6th week, fish were challenged intraperitoneally with A. hydrophila and survival percentage was recorded over 10 days post-challenge. Studied immunological parameters viz. serum lysozyme, phagocytosis, serum total protein, respiratory bursts, serum IgM levels, superoxide dismutase and alternative complement pathway activities were significantly enhanced (P < 0.05) in fish groups fed with B. aerophilus KADR3 supplemented diets, with the highest values were observed in DII (108 cfu g−1) fed group. Further, B. aerophilus supplementation at 108 cfu g−1 exhibited highest post-challenge survivability i.e. 72.83%, followed by DIII (64.19%) and DI (41.97%). Our results collectively suggest that B. aerophilus can potentially be used as probiotic strain in aquaculture to enhance the immunity and disease resistance with an optimal dietary supplementation of 108 cfu g−1.